Influenza A virus causes one of the most wide-spread infections in humans. It is also a possible bioterrorism agent. In a typical year, it infects 10-20% of the population in the US, causing up to 40,000 deaths. During the 1918 influenza virus pandemic, over 40 million people died worldwide. The threat of a new pandemic persists because the number of avian influenza outbreaks and deaths is growing and the existing vaccines have numerous drawbacks 1) current vaccines are of limited efficacy in high-risk groups, such as infants and elderly, 2) the influenza viruses they target are determined by "best guess" procedures using recent prevalent strains, 3) the time required for reformulation and large-scale production is relatively long. The four approved antiviral drugs also have limited efficacy due to severe side effects, concern about compliance, and selection for resistance viruses. For these reasons, the National Institutes of Health has designated influenza infection research, which includes therapeutics, as a priority area for biodefense research. RNA interference (RNAi) is a process by which double-stranded RNA directs sequence-specific degradation of messenger RNA in animal and plant cells. RNAi appears to be ideal for interfering with influenza virus infection because 1) short interfering (si) RNA specific for influenza A virus have been shown to potently inhibit virus production (including H1, H5 and H7 virus) in cultured cells, embryonated chicken eggs, and mice, 2) combinations of two or more siRNA specific for conserved region of flu genome will prevent the emergence of resistant virus; 3) RNAi process has less requirement for host immune function. To develop most potent siRNAs as prophylaxis and therapy of influenza virus infection in humans, this phase I application proposes i) to identify most potent influenza-specific siRNAs, ii) to evaluate the capability of siRNAs to silence the target genes with mismatches derived from different virus strains, iii) to identify the best combinations of influenza-specific siRNAs, iv) to identify polymer carriers for most efficient siRNA delivery into airway epithelial cells where influenza virus infection normally occurs, v) to identify lipid carriers for most efficient siRNA delivery, vi) to evaluate efficacies of siRNAs as prophylaxis and therapy of influenza infection in mice. The last aim will be continued in phase II of the project and the lead siRNA carriers will be further modified for efficacy improvement and will be studied for their toxicology and pharmacology. [unreadable] [unreadable]